Development of tibulizumab, a tetravalent bispecific antibody targeting BAFF and IL-17A for the treatment of autoimmune disease.

We describe a bispecific dual-antagonist antibody against human B cell activating factor (BAFF) and interleukin 17A (IL-17). An anti-IL-17 single-chain variable fragment (scFv) derived from ixekizumab (Taltz®) was fused via a glycine-rich linker to anti-BAFF tabalumab. The IgG-scFv bound both BAFF and IL-17 simultaneously with identical stoichiometry as the parental mAbs. Stability studies of the initial IgG-scFv revealed chemical degradation and aggregation not observed in either parental antibody. The anti-IL-17 scFv showed a high melting temperature (Tm) by differential scanning calorimetry (73.1°C), but also concentration-dependent, initially reversible, protein self-association. To engineer scFv stability, three parallel approaches were taken: labile complementary-determining region (CDR) residues were replaced by stable, affinity-neutral amino acids, CDR charge distribution was balanced, and a H44-L100 interface disulfide bond was introduced. The Tm of the disulfide-stabilized scFv was largely unperturbed, yet it remained monodispersed at high protein concentration. Fluorescent dye binding titrations indicated reduced solvent exposure of hydrophobic residues and decreased proteolytic susceptibility was observed, both indicative of enhanced conformational stability. Superimposition of the H44-L100 scFv (PDB id: 6NOU) and ixekizumab antigen-binding fragment (PDB id: 6NOV) crystal structures revealed nearly identical orientation of the frameworks and CDR loops. The stabilized bispecific molecule LY3090106 (tibulizumab) potently antagonized both BAFF and IL-17 in cell-based and in vivo mouse models. In cynomolgus monkey, it suppressed B cell development and survival and remained functionally intact in circulation, with a prolonged half-life. In summary, we engineered a potent bispecific antibody targeting two key cytokines involved in human autoimmunity amenable to clinical development.

Tissue disposition of 2'-O-(2-methoxy) ethyl modified antisense oligonucleotides in monkeys.

This study examined the plasma pharmacokinetics, tissue distribution, and metabolism of three second generation antisense oligonucleotides in monkeys. Three groups of monkeys were treated with 10 mg/kg of each test compound by a single 2-h intravenous infusion. Oligonucleotide concentrations were measured in plasma, tissues, and urine using capillary gel electrophoresis (CGE). HPLC-MS was used to identify the metabolite(s) of the study compounds. Plasma-concentration-time profiles after infusion for the two phosphorothioate oligonucleotides were mono-exponential, but was bi- exponential for the phosphodiester oligonucleotide. Plasma clearance for the phosphodiester oligonucleotide was four- to sevenfold higher than the two phosphorothioate oligonucleotides, which was attributed to the plasma protein binding and reduced nuclease resistance. 2'-O-(2-methoxy) ethyl (MOE) modification at both 3' and 5' ends of a phosphorothioate oligonucleotide greatly enhanced the resistance to nucleases in plasma and tissue. MOE modification only at the 3' end enhanced the resistance to nucleases in plasma, but only moderately enhanced the resistance to nucleases in tissues. Urinary excretion was a minor elimination pathway for the phosphorothioate oligonucleotide, but was a major elimination pathway for the phosphodiester oligonucleotide. The results characterize the relationships between structure and disposition and will direct future modifications for therapeutic use.